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Biomedical subjects

T Kono

Publications and source records attributed to T Kono.

At least 127 records · Page 7Linked to original sources
PubMed1997

Involvement of protein tyrosine kinase p72syk and phosphatidylinositol 3-kinase in CD2-mediated granular exocytosis in the natural killer cell line, NK3.3.

The granular exocytosis pathway is one mechanism by which NK cells and CTLs induce cytolysis of target cells. Triggering through adhesion molecules such as CD2 and LFA-1 as well as Fc gammaRIII (CD16) can invoke this pathway. CD2 is a cell surface glycoprotein present on CTLs and NK cells that plays an important role in both cellular adhesion and signal transduction. Here we report that cross-linking of CD2 as well as CD16 by immobilized Abs enhances granular exocytosis in an NK cell line, NK3.3. Herbimycin, a protein tyrosine kinase (PTK) inhibitor, or wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase (PI 3-K), inhibited completely or almost completely CD2- or CD16-mediated granular exocytosis, suggesting the involvement of protein tyrosine kinases and PI 3-K in both CD2- and CD16-mediated granular exocytosis. We also observed that cross-linking of CD2 as well as CD16 enhances p72syk tyrosine kinase activity, and this enhancement correlated well with the increased tyrosine phosphorylation of several cellular proteins, including the adapter protein Shc. Furthermore, we have observed that cross-linking of CD2 as well as CD16 enhances the PI 3-K activity associated with the tyrosine-phosphorylated cellular proteins and Shc. These results provide insight into the signaling pathways by which triggering of CD2 and CD16 on NK cells leads to cytolysis of target cells.

Androstadienes↗
PubMed1997

Transient corneal edema induced by nitric oxide synthase inhibition.

The aim of the study is to identify nitric oxide synthase (NOS) in the rabbit cornea and further investigate the physiological role of nitric oxide in the rabbit cornea. For histological identification, an immunohistochemical technique using anti-NOS monoclonal antibodies was employed. For the physiological study, we measured the corneal thickness in vivo as an indicator of corneal edema by ultrasonic pachymetry. The measurements were repeated before and after ipsilateral injections of N(G)-nitro-L-arginine methyl ester (L-NAME) or N(G)-nitro-D-arginine methyl ester (D-NAME) or 6-anilino-5,8-quinolinedione (LY-83583) with contralateral injection of vehicle (balanced salt solution) into the anterior chamber of the rabbit. We also monitored intraocular pressure (IOP) by pneumatonometry. Endothelial NOS (eNOS) immunoreactivity was demonstrated both in the corneal epithelium and the endothelium. The corneal thickness significantly increased after L-NAME or LY-83583 without significant rise of IOP, whereas no change was detected after vehicle or D-NAME. These results suggest that NO is spontaneously produced in the corneal endothelium and the NO/cyclic GMP pathway is involved in maintainance of corneal thickness.

Aminoquinolines↗
PubMed1997

Comparison of immunostainings for proliferating cell nuclear antigen and Ki-67 in human extraocular lesions.

BACKGROUND: Cell kinetic information is helpful to understand disease progression, treatment response, and prognosis of the neoplasms. To compare the usefulness and limitations of antibodies that recognize the cell cycle-associated molecules, proliferating cell nuclear antigen (PCNA) and Ki-67, we performed immunostaining in formalin-fixed, paraffin-embedded tissue sections of extraocular lesions. METHODS: Specimens were obtained from patients undergoing routine surgical procedures. Formalin-fixed, paraffin-embedded sections were stained for PCNA and Ki-67 using the monoclonal antibodies PC10 and MIB-1, respectively. Microwave oven heating for antigen retrieval was performed before immunostaining. RESULTS: In squamous cell carcinomas and basal cell carcinomas, PCNA immunostaining varied greatly. Basal cells of benign epithelial lesions showed moderate to weak PCNA staining. Strong PCNA immunoreactivity was demonstrated in foci of inflammation and germinal centers. Microwave processing enhanced the intensity of those PCNA immunostainings. As the PCNA immunostaining intensity had a great variability, the absolute numbers of PCNA-positive cells were hard to count in some tissues. High Ki-67 counts were observed in squamous cell carcinomas, foci of inflammatory cells, and germinal center cells. Basal cell carcinomas and benign lesions showed low Ki-67 counts. Every section showed clear nuclear staining in Ki-67 immunostaining. CONCLUSION: Careful consideration is required in the assessment of cell proliferation using PCNA. The immunostaining of Ki-67 may be more accurate than that of PCNA for evaluating cell proliferation in formalin-fixed, paraffin-embedded tissues.

Animals↗
PubMed1997

Localization of a G-protein-coupled inwardly rectifying K+ channel, CIR, in the rat brain.

The cellular localization of a G-protein-coupled K+ channel, CIR, in the rat brain has been demonstrated using a CIR-specific antibody, in combination with in situ hybridization. The CIR protein and messenger RNA were found in the cerebellar cortex, hippocampal formation, olfactory system, cerebral cortex, basal ganglia, several nuclei of the lower brain stem and the choroid plexus. In contrast to the messenger RNA, which was concentrated in the cell soma, the CIR protein was found in a subset of nerve fibers and, in other cases, in axon terminals. In the cerebellar cortex and hippocampus, the CIR protein was concentrated in the axon terminals of basket cells which are known to be GABAergic interneurons. This discrepancy between the distribution of protein and messenger RNA was observed in the substantia nigra, the interpeduncular, trigeminal, hypoglossal, oculomotor and red nuclei of the lower brain stem, and the tufted and mitral cells of the olfactory bulb. These observations suggested the translocation of the CIR protein into the nerve fibers following synthesis in the cell soma. Furthermore, its specific neuronal localization, especially in GABAergic interneurons, suggested the importance of CIR in synaptic transmission in neuronal systems.

Animals↗
PubMed1997

Central thyrotropin-releasing hormone stimulates hepatic DNA synthesis in rats.

Central neuropeptides play a role as physiological regulators in the autonomic nervous system. One of these neuropeptides, thyrotropin-releasing hormone (TRH), is distributed throughout the central nervous system (CNS) and acts as a neurotransmitter to regulate gastric functions through the vagus nerve. However, the autonomic nervous system is also involved in hepatic regeneration, but the effect of TRH is unknown. Therefore, the CNS's effect of TRH on hepatic DNA synthesis was studied in rats. Hepatic DNA synthesis was assessed by [Methyl-3H]thymidine incorporation 6, 12, 24, 48, and 72 hours after intracisternal injection of the TRH analog, RX 77368 (1, 5, 10, and 100 ng), and by 5-bromo-2'-deoxyuridine (BrdU) labeling of the liver section. Hepatic DNA synthesis was stimulated by intracisternal TRH analog (10 ng), with a peak response at 24 hours after peptide injection, and returned to baseline by 72 hours. This stimulatory effect by central TRH analog on hepatic DNA synthesis was dose-related, ranging from 1 ng to 10 ng (dpm/microg DNA at 24 hours [mean +/- SE]: saline, 95 +/- 6; 1 ng, 114 +/- 14; 5 ng, 318 +/- 57; 10 ng, 693 +/- 78; 100 ng, 710 +/- 135). Hepatocytes were randomly labeled by BrdU 24 hours after intracisternal TRH analog (10 ng). Intravenous TRH analog (10 ng) did not influence hepatic DNA synthesis. The stimulatory effect of TRH analog was blocked by hepatic branch vagotomy and atropine, but not by hepatic sympathectomy, 6-hydroxydopamine, insulin antibody, or hypophysectomy. These results indicate that TRH acts in the CNS to stimulate hepatic DNA synthesis through vagal and cholinergic mechanisms, and that TRH may be the chemical messenger involved in brain regulation of hepatic proliferation.

Animals↗
PubMed1997

Stage-dependent development of rat 1-cell embryos in a chemically defined medium after fertilization in vivo and in vitro.

The present study was conducted to examine the development of rat 1-cell embryos cultured in a chemically defined medium (mR1ECM) soon after penetration of eggs in vivo or in vitro. When eggs were recovered from naturally mated females at 0600-0900 h on the following day, no penetrated eggs with male pronuclei were observed. However, the percentage of pronuclear eggs had increased significantly at 1000-1100 h (17%) and 1200-1300 h (65%). When penetrated eggs recovered at 0600-0900 h were cultured in mR1ECM, blastocyst formation (22-46%) was significantly less frequent than in those (79-93%) recovered at 1000-1300 h. After collection at 0600-0800 h, preculture of penetrated eggs in modified Krebs-Ringer bicarbonate solution (mKRB) for up to 1200-1300 h improved their development to the blastocyst stage from 42% to 78%. Pronuclei were formed in almost all (98-100%) of the penetrated eggs examined after 6-20 h of insemination in mKRB. When penetrated eggs were transferred from mKRB into mR1ECM after 4-30 h of insemination, the percentages (47-64%) of blastocyst formation were significantly higher than those (1-20%) for eggs transferred after 1-3 h or after 40 h of insemination. When a total of 70 morulae or early blastocysts that had been produced by in vitro fertilization and developed in vitro were transferred to 7 pseudopregnant rats, 5 recipients into which 48 embryos had been transferred maintained their pregnancies and 25% of the embryos developed into late fetuses or pups. The results of the present study indicate that rat 1-cell embryos recovered from oviducts before pronuclear formation, or produced by in vitro fertilization, can develop to the blastocyst stage in vitro and that one or more factors in mKRB are necessary to maintain their development in mR1ECM.

Animals↗
PubMed1997

Nuclear transfer and reprogramming.

Nuclear transfer techniques for mammalian embryos have been developed in the last decade. Embryonic nuclei from advanced stages of preimplantation development can be fully reprogrammed and the totipotency is restored when nuclei are transferred into ooplasts. Transfer of nuclei after gene expression from the embryonic genome has started does not appear to restrict the reprogramming of these nuclei. The principles of nuclear transfer are outlined with respect to nuclear remodelling, nucleocytoplasmic interactions and effects of the cell cycle. However, the molecular mechanisms involved in reprogramming donor nuclei remain unknown. It is proposed that epigenetic DNA modification, such as DNA methylation that regulates gene expression, is related to the reprogramming of transplanted nuclei.

Animals↗
PubMed1997

Host defensive, immunological, and microbiological observations of an early-onset periodontitis patient with virus-associated hemophagocytic syndrome.

Virus-associated hemophagocytic syndrome (VAHS) is a disorder characterized by benign generalized histiocytic proliferation and marked hemophagocytosis associated with systemic viral infection. An immunodeficiency which includes an extremely decreased leukocyte and platelet count together with abnormalities in the CD4/CD8 ratio are the most common features of VAHS. Here we report an early-onset periodontitis (EOP) patient with VAHS from the standpoint of host-parasite interaction to understand the effect of this systemic disorder which might possibly influence susceptibility to periodontal disease. The patient is a 16-year-old Japanese male clinically diagnosed as having generalized EOP with slight gingival inflammation and moderate bone loss. This patient manifested VAHS at 3 years of age, and then had an unusual 4 recurrences (at 5, 7, 11, and 14 years old). Laboratory tests conducted include: 1) complete blood analyses: 2) peripheral neutrophil functions (chemotaxis, phagocytosis, superoxide production, and adherence); 3) peripheral lymphocyte subpopulations and functions, T-cell proliferative activity and productivity of cytokines (interleukin-2 [IL-2], interferon gamma [IFN-gamma], and tumor necrosis factor alpha [TNF-alpha]); 4) serum cytokine levels (IL-1 beta, IL-2, soluble IL-2 receptor [sIL-2R], IL-4, IL-6, IFN-gamma, and TNF-alpha; 5) serum immunoglobulin G (IgG) antibody titers against periodontopathic bacteria; 6) serological human leukocyte antigen (HLA) typing; and 7) determination of bacterial flora of the periodontal pockets. The results indicated that the patient's neutrophil chemotaxis and random migration were below the normal range. In lymphocyte examinations, T-cell proliferative activity, IL-2, and IFN-gamma productivity were elevated. Serum IFN-gamma level was also significantly higher than normal range. No specific periodontopathic bacteria were predominant in the periodontal pockets, however, the serum IgG titer against Porphyromonas gingivalis was elevated throughout the examination period. It is suggested that VAHS might be a possible risk factor for periodontal disease, and hence may serve as a model in understanding the role of host defense mechanisms in the establishment of inflammatory periodontal disease.

Adolescent↗
PubMed1997

[Evaluation of expected AUC of carboplatin (CBDCA) predicted by Chatelut's formula in comparison with Calvert's].

Several studies have shown that carboplatin AUC seems to correlate with therapeutic outcome and hematologic toxicity. Calvert's formula predicts carboplatin AUC using GFR estimated by the 51Cr-EDTA method. It is very complicated to manage radioactive agents, so we use 24-hour creatinine clearance before carboplatin administration as GFR in Calvert's. Recently, Chatelut reported a formula to predict carboplatin AUC using serum creatinine as a variable parameter of renal function as well as age, sex and body weight. We measured the AUC of free Pt by serial sampling in 14 ovarian cancer patients who received carboplatin by one-hour infusion in the initial treatment course. We evaluated the two predictive formulas with measured AUC. Calvert's predicted AUC showed a linear correlation with actual AUC (r = 0.768, p = 0.0013). But using Chatelut's formula, the predicted AUC had a better correlation with actual AUC (r = 0.833, p = 0.0002). Actual AUC correlated inversely with nadir PLT counts (r = -0.707, p = 0.0047). In conclusion, Chatelut's prediction had a better correlation with the actual AUC of carboplatin and nadir of thrombocyte count than Calvert's. Chatelut's formula was thus more useful to predict the carboplatin AUC than Calvert's when 24-hour creatinine clearance was used as GFR.

Adult↗
PubMed1996

Production of identical sextuplet mice by transferring metaphase nuclei from four-cell embryos.

Mouse clones were produced by serial nuclear transfer commencing with the transfer of four-cell nuclei at metaphase into unfertilized ooplasts. The donor four-cell-stage nuclei were synchronized in metaphase with nocodazole. The oocytes receiving a four-cell nucleus at metaphase formed two nuclei after artificial activation and inhibition of cytokinesis with cytochalasin B. To obtain embryos with diploid sets of chromosomes, nuclei from each reconstructed embryo were transferred individually into separate enucleated fertilized one-cell embryos, thus doubling the number of identical embryos. This procedure produced a high frequency of development of reconstructed embryos to the blastocyst stage. Of 11 sets of identical embryos produced by serial nuclear transplantation, 83% developed into blastocysts, including three sets of identical septuplet blastocysts. After transfer to recipient mice, a total of 25 (57%) live young were obtained, which included one set of identical sextuplet and two sets of identical quadruplet mice.

Animals↗
PubMed1996

Circadian variations of onset of acute myocardial infarction and efficacy of thrombolytic therapy.

OBJECTIVES: The present study investigated whether the onset of acute myocardial infarction and resistance to thrombolysis have similar circadian variations. BACKGROUND: Circadian variations of the onset of acute myocardial infarction and resistance to thrombolysis in the early morning have been reported. Some studies have also reported a secondary peak incidence in late evening; however, it is not known whether the resistance to thrombolysis has a similar circadian variation in these patients. METHODS: Six hundred eight Japanese patients with an acute myocardial infarction were the subjects of the study. Two hundred forty-four of the 608 patients were treated with thrombolysis within 12 h of the onset of symptoms. One hundred thirteen patients received urokinase, and 131 patients received tissue-type plasminogen activator (t-PA) over 60 min. Patency of the infarct-related artery, the primary end point of the study, was evaluated at 60 min after the initiation of thrombolytic therapy, and Thrombolysis in Myocardial Infarction (TIMI) grade 0, 1 or 2 was defined as resistant to thrombolysis. RESULTS: The onset of acute myocardial infarction and resistance to thrombolysis showed circadian variations with early morning and late evening peaks (p<0.001 and p<0.05, respectively). These circadian patterns showed similar distributions as evaluated with Spearman's method (r=0.70, p<0.05), although resistance to thrombolysis showed a phase difference of about 2 h earlier than the infarction incidence. The circadian variation of the resistance to thrombolysis was independent of the types of thrombolytic agents (urokinase or t-PA). CONCLUSIONS: These findings suggest that adjustment of treatment based on the time of the onset of symptoms may be warranted for the patients with acute myocardial infarction.

Aged↗
PubMed1996

Phorbol-ester-stimulated human lymphoid cell lines produce a plasminogen activator modulator inducing cell-bound urokinase-type plasminogen activator in malignant tumor cell lines.

The importance of cell-associated plasminogen activation in tumor invasion and metastasis is becoming increasingly evident. To clarify the modulators of cell-associated plasminogen activation in malignant states, we have recently established an assay system utilizing endogenous plasminogen activators on the cell surface. In the present study using the assay system, we found that the conditioned medium from phorbol 12-myristate 13-acetate (PMA)-stimulated human lymphoid cell lines, HUT 78 and Raji, strongly enhanced plasminogen activator (PA) activity on the surface of human malignant tumor cell lines (WI-38 VAI3 2RA, A431, A549 and HT-1080). The enhancing effect was inhibited by the addition of actinomycin D. By gel filtration, the active substances in PMA-stimulated HUT 78- and Raji-conditioned media were eluted in similar fractions corresponding to molecular weights of 60 to 80 kDa. The active substance was heat-labile. The enhanced PA activities were completely inhibited by anti-urokinase-type plasminogen activator (uPA) IgG. Moreover, the active substance was found to increase in cell-bound uPA antigen. These findings suggest that a population of activated lymphocytes produces a plasminogen activator modulator that induces uPA on the surface of malignant tumor cells.

Cell Line↗
PubMed1996

Comparison of continuous versus pulsed CO2 and Nd:YAG laser-induced pulmonary parenchymal lung injury in a rabbit model.

BACKGROUND AND OBJECTIVE: Laser techniques have recently been described for treatment of patients with emphysema and bullous lung disease. Laser exposure of the pulmonary parenchyma during these procedures is complicated by laser-induced lung injury with substantial associated morbidity. Animal investigations are needed to develop methods that reduce lung injury. We hypothesized that the depth of injury could be reduced by pulsing laser exposures, with the goal of limiting thermal effects to more superficial tissue levels. In this study, we compared acute and chronic histologic injury resulting from pulsed- versus continuous-mode CO2 and Nd:YAG laser pulmonary parenchymal exposures in rabbits. STUDY DESIGN/MATERIALS AND METHODS: A total of 40 New Zealand White (NZW) rabbits underwent thoracotomy followed by exposure with CO2 laser (n = 10 continuous vs. n = 10 pulsed at 250 Hz with duty cycle 0.15 ms) or ND:YAG laser (n = 10 continuous vs. n = 10 pulsed at 10 Hz with duty cycle 0.10 sec) to the visceral pleural surface using 1 minute of laser exposure (5 watts, defocused to 70 W/cm2 power density) to the exposed lung surface. Rabbits were sacrificed at 4 and 21 days post-injury, and lungs were examined histologically. RESULTS: CO2 and Nd:YAG laser treatment resulted in substantial pulmonary parenchymal injury. While CO2 laser-induced damage was distinct from Nd:YAG histologically, pulsed-mode laser exposures did not reduce lung injury for either laser. Acute edema occurred to depths of 1180 +/- 338 microns for continuous-mode CO2 laser exposures compared to 1,340 +/- 430 microns in pulsed mode (p = .77). For Nd:YAG laser exposure, acute edema depth was 750 +/- 748 microns continuous versus 1120 +/- 367 microns pulsed mode (p = .65). Chronic lung fibrosis depth was 450 +/- 164 microns for CO2 continuous mode compared to 575 +/- 170 microns in pulsed mode (p = .61). Lung fibrosis depth for Nd:YAG was 550 +/- 96 microns, continuous versus 484 +/- 180 microns pulsed mode (p = .76). CONCLUSION: The similarity in injury between pulsed- and continuous-mode exposures suggests that thermal relaxation times are long relative to the selected pulse frequencies in intact living rabbit lungs. Alternatively, brief high-energy pulsations may increase focal temperatures with a tendency to increase injury depth relative to the penetration of the laser light. Thus, pulsed laser modes in these settings appear to be ineffective in reducing laser-induced lung injury in clinical settings.

Animals↗
PubMed1996

Butylated hydroxyanisole blocks the inhibitory effects of tumor necrosis factor-alpha on collagen production in human dermal fibroblasts.

Tumor necrosis factor-alpha (TNF-alpha) has been demonstrated to selectively decrease the production of type I and type III collagens in human dermal fibroblasts. The effects of the commonly used food antioxidants, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), alpha-tocopherol, propyl gallate, superoxide dismutase (SOD), and catalase on TNF-alpha-induced growth enhancement and collagen metabolism were evaluated in the present study. BHA at concentrations of both 5 x 10(-5) and 10(-4) M inhibited cell proliferation and DNA synthesis induced by 10 ng/ml TNF-alpha in human dermal fibroblasts, while other antioxidants had minimal effects. Further, BHA (5 x 10(-5) M and 10(-4) M) significantly blocked TNF-alpha-induced decreases in collagen synthesis. These results suggest that antioxidants such as BHA may be involved in the modulation of collagen synthesis by TNF-alpha in human dermal fibroblasts.

Antioxidants↗
PubMed1996

Aortic counterpulsation may improve late patency of the occluded coronary artery in patients with early failure of thrombolytic therapy.

OBJECTIVES: Using a prospective, randomized design, we tested our hypothesis that the augmentation of diastolic pressure by intraaortic balloon counterpulsation (IABP) would improve the late patency of the occluded coronary artery in patients with early failure of thrombolytic therapy. BACKGROUND: Rescue angioplasty is often performed in patients in whom thrombolysis has failed, although 30% to 60% of the infarct-related arteries that are closed early after thrombolytic therapy will open later with conservative therapy. METHODS: The study included 45 patients in whom thrombolysis had failed, despite treatment with intravenous tissue-type plasminogen activator (alteplase 0.75 mg/kg body weight) delivered over 60 min within 12 h of the onset of symptoms. All patients underwent coronary angiography 60 min after initiation of thrombolytic therapy (baseline), and Thrombolysis in Myocardial Infarction (TIMI) grade 0, 1 or 2 flow was defined as failed thrombolysis. The patients were randomized to groups receiving IABP for 48 h (n = 23) or conservative therapy (n = 22, control subjects) at the end of cardiac catheterization. The late patency of the infarct-related artery, the primary end point of the study, was evaluated 3 weeks after myocardial infarction. Stenosis of the infarct-related artery was measured using a computer-assisted quantitative angiographic system in blinded manner. Data are expressed as mean value +/- SEM. RESULTS: There was no difference with regard to the baseline value for TIMI flow grade between the groups. However, 3 weeks after myocardial infarction, the patients treated with IABP had a significantly higher frequency of TIMI flow grade 3, lower residual percent stenosis and larger minimal lumen diameter of the infarct-related artery than did the control subjects (74% vs. 32%, p < 0.05; 42 +/- 5% vs. 68 +/- 6%, p < 0.01; and 1.6 +/- 0.1 vs. 0.9 +/- 0.2 mm, p < 0.01, respectively). CONCLUSIONS: These findings suggest that in patients with early failure of thrombolytic therapy, IABP may improve late patency of the occluded coronary artery, probably due to augmented perfusion pressure.

Constriction, Pathologic↗
PubMed1996

Epigenetic modifications during oocyte growth correlates with extended parthenogenetic development in the mouse.

In mammals, the maternal and paternal genomes are required for embryonic development. This is due to genomic imprinting which leads to the expression or repression of genes solely on the basis of the parent from which they were inherited. As a result, parthenogenetic embryos die before day 10 of gestation and show limited development of extra-embryonic membranes. Maternal imprinting is established during oogenesis and is associated with allele specific modifications in DNA methylation. We have investigated epigenetic modifications during oocyte growth using nuclear transfer techniques to produce mature oocytes with maternal chromatin derived from non-growing oocytes. Parthenogenetic activation of such oocytes leads to the development of normal size fetuses with a well developed placenta on day 13.5 of gestation; three days further than previously recorded for parthenogenetic development. In contrast, after fertilization, only one embryo was recovered on 9.5 days of gestation. Further, in these embryos we investigated the well characterized methylation pattern of the maternally expressed insulin-like growth factor II receptor gene (Igf2r) and found that the pattern of methylation was indeed different to that of fertilized control embryos. Thus, the embryonic phenotypes observed here correlate with changes in epigenetic events that normally occur during oocyte growth.

Animals↗
PubMed1996

A cell cycle-associated change in Ca2+ releasing activity leads to the generation of Ca2+ transients in mouse embryos during the first mitotic division.

We have used Ca2+-sensitive fluorescent dyes to monitor intracellular Ca2+ during mitosis in one-cell mouse embryos. We find that fertilized embryos generate Ca2+ transients at nuclear envelope breakdown (NEBD) and during mitosis. In addition, fertilized embryos arrested in metaphase using colcemid continue to generate Ca2+ transients. In contrast, parthenogenetic embryos produced by a 2-h exposure to strontium containing medium do not generate detectable Ca2+ transients at NEBD or in mitosis. However, when parthenogenetic embryos are cultured continuously in strontium containing medium Ca2+ transients are detected in mitosis but not in interphase. This suggests that mitotic Ca2+ transients are detected in the presence of an appropriate stimulus such as fertilization or strontium. The Ca2+ transient detected in fertilized embryos is not necessary for inducing NEBD since parthenogenetic embryos undergo nuclear envelope breakdown (NEBD). Also the first sign that NEBD is imminent occurs several minutes before the Ca2+ transient. The Ca2+ transient at NEBD appears to be associated with the nucleus since nuclear transfer experiments show that the presence of a karyoplast from a fertilized embryo is essential. Finally, we show that the intracellular Ca2+ chelator Bapta inhibits NEBD in fertilized and parthenogenetic embryos in a dose-dependent manner. These studies show that during mitosis there is an endogenous increase in Ca2+ releasing activity that leads to the generation of Ca2+ transients specifically during mitosis. The ability of Ca2+ buffers to inhibit NEBD regardless of the presence of global Ca2+ transients suggests that the underlying cell cycle-associated Ca2+ releasing activity may take the form of localized Ca2+ transients.

Animals↗